The long-term goals of this research are to understand the role of proteases in the development of renal diseases, and particularly in the pathophysiology of DN. Recent segregation and linkage analyses of Pima Indians have identified a region on chromosome 18 indicated to have a major effect on the prevalence of DN. The only candidate gene identified in this region was the structural gene for meprin-b, a subunit of a membrane metalloprotease localized to the brush border membranes of renal proximal tubules. Meprin-b is capable of cleaving a variety of bioactive peptides and proteins, including glucagon, insulin B chain, parathyroid hormone, gastrin, cholecystokinin, protein kinase A, gelatin, and collagen. The focus of this application is to test the hypothesis that variation in the meprin-b gene (structural or regulatory regions) in Pima Indians results in the variable expression, targeting, activity, or stability of the meprin-b protein, and that this is related to the susceptibility to DN. In addition to analyzing DNA from Pima Indians with advanced DN or with long-standing diabetes without nephropathy, genetically modified mice that overexpress or are null for the meprin-b gene will be developed and examined for kidney function and susceptibility to DN. The Specific Aims of the application are to: 1) analyze the meprin-b gene from Pima Indians that developed DN and unaffected individuals; 2) determine whether mutations identified in patients with nephropathy affect biosynthesis, localization, stability, or activity or meprin-b in transfection experiments, and determine whether there are abnormalities in immunochemical localization of meprin-b in kidney cortex samples from-individuals with DN; 3) overexpress the meprin-b gene in transgenic mice, and determine the effects on kidney function and susceptibility to DN induced by streptozocin or in a genetic model of type 2 diabetes; and 4) investigate the meprin-b knock-out mouse for kidney function and susceptibility to DN. These studies will establish whether the meprin-b gene is associated with susceptibility to DN in humans, and provide animal models to determine the role of this proteinase in normal kidney and in susceptibility to renal disease.